Navigational mechanisms of migrating monarch butterflies.

Abstract

Recent studies of the iconic fall migration of monarch butterflies have illuminated the mechanisms behind their southward navigation while using a time-compensated sun compass. Skylight cues, such as the sun itself and polarized light, are processed through both eyes and are probably integrated in the brain's central complex, the presumed site of the sun compass. Time compensation is provided by circadian clocks that have a distinctive molecular mechanism and that reside in the antennae. Monarchs might also use a magnetic compass because they possess two cryptochromes that have the molecular capability for light-dependent magnetoreception. Multiple genomic approaches are now being used with the aim of identifying navigation genes. Monarch butterflies are thus emerging as an excellent model organism in which to study the molecular and neural basis of long-distance migration.

A. A migrating monarch in flight. Reproduced, with permission, from Monarch Watch (www.monarchwatch.org).B. Migration south. Eastern North American monarch butterflies undergo a long-distance fall migration (red arrows east of the Rocky Mountains) to a restricted destination in central Mexico (yellow oval). The population of monarch butterflies west of the Rocky Mountains also undergo a fall migration, overwintering in protected roosts along the Pacific Coast (red arrows west of the Rocky Mountains), but the distances traveled are far less than those of the eastern population. In addition, the western range of overwintering areas is not as concentrated as that in the east. Because of the distances traveled and the nature of the focal stopping point, most navigational research has centered on the eastern North American migration.C. Journey north. The Eastern North American migrants remain at the overwintering areas in Mexico until spring, when the same butterflies reproduce and disperse northward to lay fertilized eggs on newly emerged milkweed in the southern United States (red arrows). Successive generations of spring and summer monarchs in the east and the west re-populate the home range (black arrows).

Essential role for a functioning circadian clock in sun compass orientation

A. Flight simulator. An individual monarch butterfly is affixed to a directional recording device using a short length of wire. It may fly and rotate freely, but cannot move vertically or horizontally. The tethered butterfly is suspended in a plastic barrel, excluding buildings and trees from view. During flight, a video recorder monitors the butterfly and the directional recording device sends orientation information to a computer. Adapted, with permission, from Ref. [].B. Flight orientation of migrants housed under different lighting conditions. Large circle represents the 360° of possible direction (0° is north); small circles represent individual flight orientation of migrants. The arrow indicates mean vector, and the length of the arrow represents strength (r value). The butterflies in the top graph were housed under normal light/dark conditions in the lab, with simulated sunup at 7 AM and simulated sundown at 7PM. Another group of butterflies (bottom graph) were clock-shifted by 6 hours, with simulated sunup at 1 AM and simulated sundown at 1 PM. So when tested in the flight simulator at ~10 AM, the normal housed butterflies perceived it to be 10 AM and appropriately oriented to the southwest, with a mean vector (α) of 233°, while the clock-shifted butterflies perceived it to be 4 PM and thereby shifted their oriented counterclockwise by 115° to the southeast; the sun’s azimuth varied from 16° to 22° per hour over the course of study, which would result in a 6-hour shift of between 96° and 132°. This clock-shift experiment shows the importance of the timing of the circadian clock for proper flight orientation. Each butterfly used flew continuously for 10 min. Adapted, with permission, from Ref. [].

A. Molecular clockwork model. The main gear of the clock mechanism is an autoregulatory transcription feedback loop in which CLOCK (CLK) and CYCLE (CYC) heterodimers drive the transcription of the period (per), timeless (tim), and cryptochrome2 (cry2) genes through E box enhancer elements. TIM (T), PER (P), and CRY2 (C2) are translated and form complexes in the cytoplasm; 24 hours later CRY2 is shuttled into the nucleus to inhibit CLK:CYC-mediated transcription. PER is progressively phosphorylated and likely helps translocate CRY2 into the nucleus. CRYPTOCHROME1 (CRY1, C1) is a circadian photoreceptor which, upon light exposure (lightning bolt) causes TIM degradation, allowing light to gain access to the central clock mechanism for photic entrainment. The monarch clockwork, which has both Drosophila-like and mammalian-like aspects, was formulated from a combination of in vitro and in vivo approaches, including the use of the monarch DpN1 cell line, which contains a light-driven diurnal clock, and the use of Drosophila transgenesis, to augment the differential clockwork functions of the monarch CRY proteins []. Adapted, with permission, from Ref. [].B. Schematic representation of brain clocks and circuits in the monarch butterfly brain. Cells expressing TIM, PER, CRY1 or CRY2 are highlighted in blue [, ]. In these areas the clock proteins partially colocalize. All four clock proteins are co-localized in two of the four cells in the pars lateralis (PL) on each side of the brain; these four cells in total are the bona fide circadian clock cells in brain [, ]. CRY1-positive cell bodies and projections are represented by green dots and green lines, respectively. Projections from the dorsal rim area photoreceptors are indicated by the dotted orange lines. Neurons and fibers expressing exclusively CRY2 are represented in red and within the central body are shown as red circles and red hatching. PI, pars intercerebralis; PL, pars lateralis; CC, central complex; LA, lamina; ME, medulla; LO, lobula. Adapted, with permission, from Ref. [].

Model of the components and potential circuitry involved in the time-compensated sun compass mechanism

(left) Skylight input to the eye: Skylight cues provide directional information that is sensed by the eye and ultimately integrated into the central complex (CC), the presumed site of the sun compass. The dorsal rim area of the eye senses the angle of plane-polarized, ultraviolet light (small violet circle with crosshatches), while the main retina senses color gradients (small filled violet circle, small blue circle, and small green circle) or senses the sun itself (i.e., does not discriminate between the three colors). Multicircuit pathways, yet to be defined, connect eye-sensed skylight information to the central complex (dashed black and grey lines with question mark).(right) Clock entrainment: Skylight in the blue range directly entrains (synchronizes) circadian clocks in the antennae and brain to the 24-hour day through direct light action on CRYPTOCHROME1. An as yet unknown neural pathway connects antennal clocks to the central complex (thick dashed red line with question mark); this is a major pathway providing timing information to the central complex. A neural pathway that connects clock cells in the pars lateralis (PL; blue spots) area of the brain to the central complex (thin dashed red line) is likely to exist; this may provide a minor pathway for timing sun compass orientation. (below) Signal integration: Information from the sun compass and circadian clock is integrated in the central complex itself or in its output pathways. Central complex output pathways communicate with the motor system to ultimately control continuous flight in the southwestly direction.